Kwatra_1986_J.Biol.Chem_261_12429

We have tested the possibility that regulation of cardiac muscarinic receptor function may involve receptor phosphorylation. Chick heart muscarinic receptors were purified from relatively small amounts of tissue to near homogeneity using a three-step chromatographic procedure that utilized the affinity chromatography procedure of Haga and Haga (Haga, K., and Haga, T. (1983) J. Biol. Chem. 258, 13575-13579). The purified preparations contained a single major peptide which migrated on sodium dodecyl sulfate gels with an apparent Mr of 79,000. When receptors were purified from 32P-bathed hearts, a single major phosphopeptide eluted from the affinity column and comigrated on sodium dodecyl sulfate gels with the band of stained receptor. Treatment of hearts with the agonist carbachol led to marked increases (10-12-fold) in the phosphorylation of the receptor. The results show that the muscarinic receptor is a phosphoprotein in cardiac tissue and that treatment with a receptor agonist regulates its phosphorylation in the intact cell.